The present work aims to develop a growth medium to render a wild-type strain of Saccharomyces cerevisiae permeable to the antifungal drug Brefeldin A. In the current study, a synthetic medium containing 0.1% L-proline and supplemented with 3.0 × 10 −3 % SDS is employed. When Brefeldin A is added to this medium, a wild-type strain shows increased growth sensitivity and a diminished transport of the amino acid L-leucine. Since Brefeldin A exerts its effect on the endoplasmic reticulum and the Golgi apparatus, the medium permits the study of the drug effect on the intracellular traffic of L-leucine permeases. INTRODUCTIONBefore their delivery to the plasma membrane (PM), the different permeases involved in amino acid transport, like most of the membrane proteins, enter the membrane of the endoplasmic reticulum (ER). They then proceed through the protein secretory pathway of the ER, via the Golgi complex (GC) and exocytic vesicles, until they finally reach the PM [1].A very useful agent for investigating permease transport through the secretory pathway is the antifungal agent, Brefeldin A (BFA), which reversibly blocks the transport of proteins from the ER to the Golgi [2,3,4]. This drug can be used to create a temporary block in transport, allowing accumulation of permeases in the ER and depletion of these permeases downstream. In addition, when the BFA block is present, loss of permease molecules from the PM through endocytosis can be studied independent of their replacement via the secretory pathway. Moreover, release of the BFA block would permit the investigation of the dynamics of replacing the permeases in the depleted membrane. Because wild-type yeast has a very low apparent permeability to BFA, previous investigations have used strains bearing the erg6 mutation that blocks the final methylation reaction in ergosterol biosynthesis. The lack of ergosterol in the PM changes the permeability properties of the membrane and renders cells sensitive to several inhibitors, including BFA and the dye, crystal violet (CV) [2]. These changes appear to be at least partly due to decreases in activity of multidrug resistance pumps such as Pdr5p [5].There are several disadvantages of using the erg6 mutation to obtain BFA sensitivity. The mutation itself causes a marked increase in permeability to sodium and lithium ions [6]. Efficiency of genetic transformation is lowered dramatically, and sexual conjugation is also greatly reduced. Moreover, transport of tryptophan is lowered substantially [7].We have developed a simple method for obtaining BFA sensitivity without requiring the introduction of erg6. Because the method requires no genetic manipulation, it can be applied to wild-type cells and to strains already bearing various mutations related to secretion, to altered amino acid transport, and to modified permease turnover. The method depends upon the use of an SDS-supplemented synthetic growth medium in which the wild-type strain MMY2 presents increased sensitivity to BFA. At appropriate concentrations, BFA inhibits grow...

Carnitine is well-known for its role in the transport of fatty acids to the mitochondrial matrix, where beta-oxidation takes place. This work describes novel functions for this compound and novel data on its pharmacokinetics.

In sepsis, reactive oxygen species (ROS) production is increased. This process takes place mainly within the electron transport chain. ROS production is part of the pathophysiology of multiple organ failure in sepsis. Succinate yields Dihydroflavine-Adenine Dinucleotide (FADH2), which enters the chain through complex II, avoiding complex I, through which electrons are lost. The aim of this work is to determine if parenteral succinate reduces systemic ROS production and improves kidney function. Rats with cecal ligation and puncture were used as model of sepsis, and 4 groups were made: Control group; Succinate group, which only received parenteral succinate; Sepsis group; and Sepsis which received parenteral succinate. Systemic ROS are measured 24 hours after the procedure. Rats subjected to cecal puncture treated with succinate had less systemic ROS than Septic untreated rats (p = 0.007), while there were no differences in creatinine levels (p = 0.07). There was no correlation between creatinine and systemic ROS levels (p = 0.3). We concluded that parenteral succinate reduces ROS levels, but it does not reduce creatinine levels. Since there is no correlation between both levels, the processes would not be related.

We focused on the participation of GAP1, BAP2, and AGP1 in L-phenylalanine transport in yeast. In order to study the physiological functions of GAP1, BAP2, and AGP1 in L-phenylalanine transport, we examined the kinetics, substrate specificity, and regulation of these systems, employing isogenic haploid strains with the respective genes disrupted individually and in combination. During the characterization of phenylalanine transport, we noted important regulatory phenomena associated with these systems. Our results show that Agp1p is the major transporter of the phenylalanine in a gap1 strain growing in synthetic media with leucine present as an inducer. In a wild type strain grown in the presence of leucine, when ammonium ion was the nitrogen source, Bap2p is the principal phenylalanine carrier.